84 results about "Insecticidal crystal proteins" patented technology

The subject invention pertains to the use of peptide fragments of cadherins (including cadherin-like proteins). The subject invention includes a cell (and use thereof) comprising a polynucleotide that expresses the peptide fragment. The subject invention includes methods of feeding the peptides to insects. In preferred embodiments, the peptides are fed to target insects together with one or more insecticidal proteins, preferably (but not limited to) B.t. Cry proteins. When used in this manner, the peptide fragment can not only enhance the apparent toxin activity of the Cry protein against the insect species that was the source of the receptor but also against other insect species. Preferably, the cadherin is a Bacillus thuringiensis (B.t.) insecticidal crystal protein (Cry) toxin receptor. Preferably, the peptide fragment is a binding domain of the receptor. In some preferred embodiments, the peptide is the binding domain nearest to the membrane proximal ectodomain. Corresponding domains are identifiable in a variety of B.t. toxin receptors.

A Bacillus thuringiensis insecticidal crystal protein DNA sequence Cry2A is designed and synthesized. Comparing it with original Cry2A DNA sequence can show that the amino acid components of DNa sequence coding protein are not changed, the application frequency of vegetable preference codon is higher and the AT sequence, reverse reduplicate sequence and indefinable subsequences are eliminated. It can be used for culturing the seed of insect-resisting transgenic plant.

The invention discloses an insect-fusion-resistant gene, coding protein, a carrier and application thereof. The insect-fusion-resistant gene is formed by using a connecting peptide coding gene to connect a nucleotide sequence of coding BT insecticidal crystal protein Cry1Ab with coding BT trophophase insecticidal protein Vip3A, the connecting peptide amino acid sequence is shown as SEQ ID NO:1. Insecticidal activity of the insect-fusion-resistant gene is improved, and insecticidal spectrum is wider. The insect-fusion-resistant gene can kill main lepidopterous insects like armyworm, Spodoptera litura, Ostrinia furnacalis, cotton bollworm and Spodoptera exigua of paddy and maize. In addition, the insect-fusion-resistant gene can effectively slow down insect resistance.

The invention belongs to the technical field of microorganism genetic engineering and discloses separation and cloning of nematicidal crystal protein gene originated from thuringiensis and biological activity of the nematicidal crystal protein gene on nematode. In the invention, a new insecticidal crystal protein gene cry1518-35 is obtained by being separated from bacterial strain of activated thuringiensis YBT-1518 of root-knot nematodes in north of China; an encoded product of the gene is a new insecticidal crystal protein Cry1518-35; the protein Cry1518-35 features high insecticidal activity on root-knot nematodes in north of China. The invention also includes the preparation of colon bacillus recombinant bacteria EMB0225 which expresses the insecticidal crystal protein Cry1518-35, the recombinant colon bacillus is preserved in CCTCC, the preservation number is CCTCC NO: M209009.

The invention discloses a Bacillus thuringiensis MB-15 strain which has a broad spectrum and high virulence on vegetable lepidoptera pests, and the preservation number of the strain is CGMCC No. 5255. The strain can form a diamond insecticidal crystal protein, contains cry1I, cry1Ac, cry2Ac and vip3Aa genes, and has high-efficiency insecticidal activity on plutella xylostella, Pieris brassicae, beet armyworm, Spodoptera litura, Helicoverpa armigera and other lepidoptera pests. The invention also relates to a preparation method of the wettable powder of strain. The wettable powder comprises the following content of main components: 40.0-50.0% of Bacillus thuringiensis MB-15 strain powder, 5.0-10.0% of a dispersant, 5.0-10.0% of a wetting agent, 0.5-1.0% of a stabilizer, 0.5-1.0% of an ultraviolet light protection agent, and 28.0-49.0% of a carrier. The wettable powder can be used to control a plurality of lepidoptera pests on vegetables, and has the characteristics of broad spectrum, high efficiency, and safety, etc.

The invention discloses an insecticidal crystal protein CrylAc enzyme linked immunosorbent detection kit. In the CrylAc protein detection of the kit, the lowest detection limit is 7.81ng / mL, the linear detection range is 62.5-500ng / mL, the linear interval fitted equation y is equal to 1.1136Ln(x)-4.164, and R2 is equal to 0.9991. No cross reaction occurs among the kit and CrylAb and Crylc proteins. The kit is suitable for qualitative detection of the CrylAc protein in transgenic plant leaves, fruits and derivatives. The kit can detect large batch of samples simultaneously, is convenient and fast, has very important realistic significance in analyzing related transgenic articles, simultaneously ensures the detection cost to be greatly reduced and has potential economic value.

The invention relates to a biological suspension pesticide prepared by compounding abamectin with bacillus thuringiensis, which comprises 88-93 percent by weight of main agent and 7-12 percent by weight of auxiliary agent, wherein the main agent is a mixture of the abamectin and the bacillus thuringiensis; the auxiliary agent comprises a dispersing agent, a thickening agent and other assistants; and the dispersing agent is polyoxyalkylene ether FS1 and urea and magnesium aluminum silicate. According to the formula, the biological suspension pesticide is prepared by the steps of: adding a bacillus thuringiensis fermentation liquor, the abamectin, the urea, the magnesium aluminum silicate and the thickening agent into a ball-milling reactive kettle, stirring at low speed, adding the FS1, and stirring at high speed; and then adding the other assistants for stirring uniformly to obtain the biological suspension pesticide. In the biological suspension pesticide, the content of the abamectin is not less than 2 percent, the content of insecticidal crystal protein of the bacillus thuringiensis is kept above 80 percent, the survival rate of the bacillus thuringiensis spore is above 90 percent, and the cotoxicity coefficient of the pesticide is more than 120. The invention is beneficial to delaying the generation of pesticide resistance of insects, and has the advantages of quick effect-taking, long valid period and low cost lower than or not more than that of the traditional similar pesticides.

The invention discloses the separation, clone and application of insecticidal crystal protein gene from Bacillus thuringiensis. In this invention a novel insecticidal crystal protein cry7Ba1 is separated from Bacillus thuringiensis Central China species YBt-978 strains. The genes code a novel insecticidal crystal protein Cry7Ba1 with insecticidal activity to Lepidoptera insects. Cry7Ba1 protein is consisted by 1,154 amino acid, and its homology with the existing published proteins is no more than 80% and the half of amino acid sequence near the protein N-terminal has no more than 50% homology with the existing published proteins. The invention also discloses the gene sequence of this novel insecticidal crystal protein and its coding protein Cry7Ba1 expressed by microorganism transformed by appropriate expression vector, qualifying it with lepidoptera insecticidal activity.

The invention relates to a hollow microcapsule, an acidic or alkaline controlled-release microcapsule and preparation methods thereof, belonging to the field of biological agents. The preparation method of the hollow microcapsule comprises the following steps of: preparing a microcapsule through a layer-by-layer self-assembly method by taking calcium carbonate as a core and selecting polyelectrolyte cation and anion with opposite charges under a preparation condition; and then removing the core to obtain the hollow microcapsule. The wall of the hollow microcapsule is formed by alternating PAH / PSS (Polypropylene Amine hydrochloride / Poly Sodium Styrenesulfonate), and the hollow microcapsule can loaded with Cry protoxins of bacillus thuringiensis insecticidal crystal proteins under an acidic condition so as to obtain a microcapsule dosage form with release controlled under an alkaline condition. The microcapsule dosage form keeps the insecticidal activity of the cry protoxins, can assist the cry protoxins to resist the influence of certain environmental factors, also enables the insecticidal proteins to be specifically released in an alkaline environment of an insect midgut and keep stable in a general environment and plays an important role in the actual application of the Bt (Bacillus thuringiensis) insecticidal proteins on pest control.

The invention relates to a preparation method of a liquid fermentation medium for bacillus thuringiensis, which includes mushroom residue preprocessing and culture medium ingredient supplementation. The preparation method disclosed by the invention includes the following steps: strong acid is utilized to degrade the cellulose and pleurotus eryngii mycelium in mushroom residue into sugar and a small amount of protein, phosphorus and trace elements as main nutrient substances contained in residue extract, which can be utilized by the bacillus thuringiensis, meanwhile, nitrogen source and inorganic salt ions are supplemented partially, the bacillus thuringiensis can grow and reproduce under the condition of maintaining appropriate pH, temperature and ventilation, so that insecticidal active substances, such as spores and insecticidal crystal protein, can be produced, and the fermentation medium is concentrated and dried to form an environment-friendly and secondary pollution-free microbial insecticide product. The preparation method not only solves the problem of mushroom residue processing and prevents the mushroom residue from polluting the environment, but also realizes the comprehensive utilization of resources, and meanwhile, the preparation method also greatly reduces the production cost of the microbial insecticide, and turns waste into treasure.

The invention discloses a method for detecting insecticidal crystal proteins Bt Cry1Ab / Ac and a special enzyme linked immunosorbent assay kit thereof. An anti-Bt Cry1Ab / Ac monoclonal antibody used by the method and the kit is secreted by a hybridoma cell strain Bt2F9, and the application number of the hybridoma cell strain in CGMCC (China General Microbiological Culture Collection Center) is CGMCC No.5162. The monoclonal antibody is obtained by simultaneously using a Bt Cry1Ac protein solution and an extracting solution of Bt Cry1Ac transgenic cotton plant leaves as a coating antigen and screening hybridoma cells through an indirect non-competitive ELISA (Enzyme-Linked Immuno Sorbent Assay) method, the monoclonal antibody has high affinity for Bt Cry1Ab / Ac protein expressed in plants, and the affinity constant is 1.03*10<9>M<-1>. The double-antibody sandwich enzyme linked immunosorbent assay kit prepared by the monoclonal antibody is used for qualitatively or quantitatively detecting proteins Bt Cry1Ab / Ac in transgenic plants such as cotton and corn, the linear detection range is 0.78-50ng / mL, and the kit has the advantages of fastness and sensitiveness.

The invention relates to activated bacillus thuringiensis HLJ-66 with Bt diamond back moth resistance and application thereof. The strain disclosed by the invention is bacillus thuringiensis HLJ-66, is called HLJ-66 for short and has the preservation number of CGMCCNo.8445, the preservation data of Nov. 8th in 2013 and the preservation address of No. 3, courtyard No.1, Beichen West Road, Chaoyang District of Beijing. The strain has toxicity for anti-Cry1Ac insecticidal crystal protein and Btk diamond back moth by toxicity biological assay screening. The strain is utilized to produce a high-efficiency broad-spectrum pesticide for preventing and treating the lepidopterous important pest of diamond back moth; and meanwhile, the strain has a good effect on resistance pests, is beneficial for preventing and treating resistance diamond back moth in the field, improves application value, is harmless for people, livestock and other animals, has no pollution to the environment and has excellent ecological benefits and excellent application and popularization prospect.

The invention relates to the technical field of agriculture microorganism genetic engineering, in particular to separation and cloning of a Helicoverpa armigera cadherin gene fragment hacad1and encoded polypeptide fragment HaCad1. By separation, the cadherin gene fragment hacad1 of 735bp is obtained from the midintestine of the Helicoverpa armigera, an encoded product thereof is the polypeptide fragment HaCad1 which can be combined with Bt toxin CrylAc10. Biological experiments indicate the polypeptide prepared in the invention has very strong enhancing insecticidal activity on insecticidal crystal protein CrylAc10 of lepidoptera insects. The invention also includes preparation and an application for recombinant Escherichia coli EMB1104 expressing the Helicoverpa armigera cadherin fragment HaCad1. The recombinant Escherichia coli EMB1104 is preserved in CCTCC, the preservation number is CCTCC NO: M209010.

The invention relates to a new bacterial strain of Bacillus thuringiensis for killing grub pest and a pest killing protein thereof. The new bacterial strain has the collection number of CCTCC (China Center For Type Culture Collection) No. M2011267. The bacterial strain and the pest killing protein provided by the invention can be used for preparing the preparation capable of preventing and controlling grub pests. The bacterial strain provided by the invention has specific insecticidal activity on the grub pests. The pest killing crystal protein provided by the invention has good prevention and control effect on main soil insects such as Holotrichia oblita and Holotrichia parallela in China.

A novel gene encoding a Coleopteran inhibitory Bacillus thuringiensis insecticidal crystal protein is disclosed. The protein, tIC851, is insecticidally active and provides plant protection from at least cotton boll weevil, Anthomomus grandis, when applied to plants in an insecticidally effective composition.

The invention discloses a microfluidic protein chip for high-through detecting genetically modified crops. The microfluidic protein chip comprises a nucleic acid chip which is synthesized on the basis of mu ParafloTM microfluidic protein chip and contains six nucleotide sequences and 6 types of antibody-oligonucleotide composite probes, thereby simultaneously detecting 6 transgenetic proteins of trans-bacillus thuringiensis Bt insecticidal crystal protein (Cry1Ac), trans-bacillus thuringiensis Bt insecticidal crystal protein (Cry1c), trans-bacillus thuringiensis Bt insecticidal crystal protein (Cry1F), trans-bacillus thuringiensis Bt insecticidal crystal protein (Cry1Ab), soybean trypsin (SBTI) and neomycin phosphate transferase II gene (NPTII). The invention further provides a transgenic assay kit which comprises a mu ParafloTM microfluidic deoxyribose nucleic acid (DNA) chip which contains six types of probes, six types of antibody-oligonucleotide composite probes, six types of detection antibodies and the like.

The invention discloses a standard molecular for specifically detecting a transgenic rice strain Kefeng No.6 and an application thereof. The standard molecule disclosed by the invention is an annular carrier which comprises a cowpea trypsin inhibitor coding gene segment shown by the 10th-101st sites of a sequence 1, a bacillus thuringiensis insecticidal crystal protein coding gene segment shown by the 256th-336th sites of the sequence 1, a rice endogenous gene GOS segment shown by the 191th-258th sites of the sequence 1 and a border sequence segment of Ti plasmid T-DNA shown by the 108th-184th sites of the sequence 1. Experiments prove that the standard molecule disclosed by the invention has high sensitivity, wide linear range, good uniformity and strong stability, and is of important significance to the qualitative and quantitative detection of the transgenic rice strain Kefeng No.6; and the lowest detection line of the standard molecule can reach 2 copies.

The invention relates to a preparation process of a live bacillus thuringiensis preparation. The preparation process comprises the steps that bacillus thuringiensis is inoculated into a fermentation medium to be cultured after being subjected to primary and secondary seed enlarge culture; the quantity and conversion rate of bacillus are increased through fed-batch of a carbon source in the fermentation process, thus obviously increasing the unit yield and reducing the unit cost; after fermentation is completed, the fermentation liquor is concentrated by using a vacuum film concentrator and the soluble synergistic substances, growth-stimulated substances and bacteriostatic active substances in the fermentation liquor can be effectively recovered; the concentrated fermentation liquor is dried and granulated by multistage low temperature spray fluidized drying equipment, the granules are sorted and the fine powder is trapped by a cyclone separator and returns to be re-granulated, thus obtaining the bacillus thuringiensis product, wherein, through biological assay, the activity of each gram of the bacillus thuringiensis powder on cotton bollworms is not lower than 80000IU / mg and the content of insecticidal crystal proteins in the product is not lower than 13%; the tail gas is treated and is discharged up to the standard. The preparation process has the advantages that the active substances in the fermentation liquor can be furthest retained; the product quality and the equipment utilization rate are high; the biological activity is better maintained; the waste gas and the wastewater are discharged up to the standard; the preparation process can be used for large-scale industrial production.

The invention relates to the field of insecticidal crystal protein, in particular to Cry2-class insecticidal crystal protein with insecticidal activity on beet armyworm, striped rice borer, Asiatic corn borer and plutella xylostella.

The invention relates to an environment-friendly insecticide for killing wax-moth larvae and eggs thereof. The environment-friendly insecticide consists of the following components in percentage by weight: 0.5%-10% of a bacillus thuringiensis bactericide, 0.2%-10% of chlorinated sodium phosphate, 5%-10% of a wetting agent and 70%-92.5% of bait, wherein the bait is an old comb. The aqueous solution of the chlorinated sodium phosphate is alkaline, so that the insecticidal activity of bacillus thuringiensis insecticidal crystal protein is obviously increased; and moreover, the prevention and control objects of the environment-friendly insecticide are widened as compared with the conventional insecticides, and the environment-friendly insecticide can kill not only larvae but also eggs, takes a shorter prevention and control time than other insecticides and thoroughly kill wax moths. The bacillus thuringiensis and the chlorinated sodium phosphate are not toxic or slightly toxic to human and animals and are low in usage, the wetting agent and the bait are harmless to bees and toxic-free to human and animals; and a cloth bag separates the insecticide from the outside, so that the insecticide cannot contaminate bee products.

The invention discloses an insecticidal crystal protein gene of bacillus thuringiensis and the application thereof; the insecticidal protein coded by a DNA sequence of the insecticidal crystal protein gene shown in SEQ ID No.1 is an amino acid sequence shown in SEQ ID No.2. The insecticidal protein coded by the gene has specific insecticidal function to orthopterous oriental migratory locust, the locust insecticide prepared by the gene has good prevention effects used for preventing and curing those locusts harmful to graminaceous crops such as grain, wheat, corn, broomcorn and pasture etc., and is safe to human and animal and plant, has low production and using cost, does not cause environmental pollution, thereby having great application value.

The invention discloses a pest-resistant fusion gene, an encoded protein, a carrier, and applications. The pest-resistant fusion gene is composed via connection of a nucleotide sequence used for coding BT insecticidal crystal protein Cry1Ab with a nucleotide sequence used for coding BT vegetative insecticidal protein Vip3A or Vip3C. The encoded protein of the pest-resistant fusion gene is high ininsecticidal activity and wider in insecticidal spectrum, is capable of killing the main lepidoptera pests of rice and corn such as armyworm, prodenia litura, corn borer, cotton bollworm, and beet armyworm, and in addition, the encoded protein is capable of delaying generation of insect resistance.

The subject invention includes methods and plants for controlling European corn borer, said plants comprising a Cry1Ab insecticidal protein and a DIG-3 insecticidal protein to delay or prevent development of resistance by the insect.

The invention belongs to the technical field of microbe genetic engineering, and discloses separation and cloning of nematicidal crystallin gene originated from thuringiensis and biological activity of the nematicidal crystal protein gene on nematode. In the invention, a new insecticidal crystallin gene cry003-148 is obtained by being separated from thuringiensis Sbt003 (the preservation number in China Center for Type Culture Collection is CCTCC NO: M2012168); an encoded product of the gene is a new insecticidal crystallin Cry003-148; and the crystalline has high insecticidal activity on root-knot nematodes in south of China. The invention also comprises preparation of thuringiensis recombinant bacteria BMB1361 which expresses the insecticidal crystallin Cry003-148, the recombinant bacteria is preserved in China Center for Type Culture Collection (CCTCC) and the preservation number is CCTCC NO: M2012057.

The subject invention includes methods and plants for controlling European corn borer, said plants comprising a CrylAb insecticidal protein and a DIG-3 insecticidal protein to delay or prevent development of resistance by the insect.

The invention discloses a fusion protein capable of efficiently resisting spodoptera frugiperda. The fusion protein is formed by fusion connection of BT insecticidal crystal protein and BT vegetativeinsecticidal protein Vip3, and the BT insecticidal crystal protein is BT insecticidal crystal protein Cry1Da or BT insecticidal crystal protein Cry1Fa. The fusion protein has higher insecticidal activity on noctuidae pests, significantly improves the insecticidal activity on lepidoptera noctuidae pests such as the spodoptera frugiperda, is beneficial to the insect-resistant protein to exert the biological activity, and alleviates the risk of pest resistance. The fusion protein can be used for simultaneously and efficiently killing main lepidoptera pests on rice, corn and soybeans such as spodoptera frugiperda, armyworms, corn borers, cotton bollworms, beet armyworms, cutworms, chilo suppressalis, pink rice borers and cnaphalocrocis medinalis.

The present invention belongs to the field of gene engineering, relates to the constitution, expression and application of fusion protein, and is especially one kind of transduction peptide-insecticidal crystal protein fusing protein and its application. The PTD research results are applied in the field of microbial insecticidal crystal protein for the first time, so as to expand new field for applying PTD and open new way to eliminating pest's resistance. The fusion protein has pesticidal activity higher than that of single insecticidal crystal. The present invention has important theoretical and practical significance in controlling harmful organism and applying PTD.